Sample recovered from a sexual assaulted victim is typically a mixture of different cell types including epithelial cells, erythrocytes, white blood cells, various vaginal flora (such as Lactobacillus), sperm cells and bacterial, viral or fungal contaminants. Samples of sexual assaults including but not limited to buccal assault or anal assault may further comprise buccal epithelial cells, buccal flora, intestinal epithelial cells, colon epithelial cells, or other bacterial cells as part of the female components. Capture and detection of sperm DNA from a sample, such as a sexual assault sample are primary requirements for applications, such as forensic applications or diagnostic applications.
Different technologies have been developed to capture and/or detect sperm DNA from a sample, which includes: DNA finger printing to distinguish male DNA using Y chromosome probes, restriction fragment length polymorphism (RFLP) or variable number tandem repeats (VNTR) to determine specific patterns of sperm DNA. Nuclear and cytoplasmic stains may also help to identify a sperm cell, however the staining method relies on the intact shape of the sperm cells, which may not be obtained when present in a sexual assault sample. These methods are complex as they require multiple steps and require significant amount of starting DNA. To achieve desired concentration of starting DNA, variety of techniques have been developed, which may include amplification of target DNA. In amplification, contaminating species may also get amplified and the amplification based detection lacks the specificity for detecting cell source of the amplified DNA. These applications are typically preceded by separation and purification of target DNA from unwanted nucleic acids and contaminants to reduce interference in downstream applications and to achieve desired result. However, the traditional purification or separation methods and the associated techniques are complex, time and labor intensive. Further often a sample is compromised or degraded after a set period of time, for example is relationship to a sperm sample, often 3 to 5 days after ejaculation.
In sperm cells, the histone proteins found in somatic cells are replaced in large part by protamine (PRM 1 and PRM 2), and allow further DNA condensation during sperm maturation. Protamine is uniquely expressed in human sperm relative to other cells in the body and serves as a highly specific factor associated with the DNA in sperm cells. The presence of protamine is highly conserved across species.
Chromatin immunoprecipitation (ChIP) is a well-established method for isolating specific DNA through affinity capture of associated proteins. Traditionally ChIP has been used for transcription factor mapping to active gene promoter regions, and more recently has been adapted for analysis of the epigenome of both DNA and associated histone proteins and their respective modifications. ChIP has been used for isolation of protamine DNA complexes, however, ChIP has never been used for purifying sperm specific DNA from a sample comprising different types of cells.
A simplified method for isolating assailant DNA, specifically DNA from sperm cells, from a sample composed of victim's cells and assailant's cells in mixed sexual assault casework samples for subsequent analysis is highly desirable.